It may be of interest to study two different intracellular pathogens in the same biological sample, for example a sample isolated from a patient or from a cell culture, or to observe one intracellular pathogen in the presence of other intracellular pathogens. However, often the presence of one such pathogen interferes with observations of other pathogens.
For example, when the effects of intracellular pathogens are studied in assays leading to general phenotypic results (“phenotypic assays”), the observables are not necessarily specific for a particular pathogen. For example in virology, a plaque observed in a cell culture cannot necessarily be attributed to a given virus if a further pathogen capable of causing such plaques is also present.
However, it is often desirable to employ general phenotypic assays. Other, more specific methods of observing pathogens suffer from different drawbacks. Often, more specific methods are not as straightforward or rapid to perform. Such other methods may also require specific reagents which impose further limitations.
Immunological assays, for example, require antibodies to be available. The utility of immunological approaches may further be limited when antibodies are cross-reactive for multiple pathogens that may be present in the sample. For some diagnostic purposes, broad cross-reactivity of antibodies to related pathogens may be desirable. On the other hand, such cross-reactivity may make it impossible to study related and co-existing pathogens independently by immunological methods.
PCR-based testing for pathogens can be more specific. However, PCR-based methods require genomic sequences to be known for each pathogen of interest. Primers must be designed and optimized for each pathogen of interest,
The requirement for specific antibodies or primers limits the flexibility of both immunological and PCR-based methods, and in particular their utility for observing pathogens when the nature of the pathogen is not known. The utility of these specific methods may moreover be limited in the case of rapidly evolving pathogens, e.g., certain viruses, because mutations may occur in epitopes recognized by antibodies or in the sequences recognized by PCR primers.
It is an object of the invention to provide further and improved methods for studying intracellular pathogens of interest in a biological sample independently of other intracellular pathogens.